Several groups have demonstrated the ability to reprogram human fibroblasts to induced pluripotent stem cells (iPSCs) following transduction with Oct-4 together with other factors (Takahashi et al., 2007; Takahashi and Yamanaka, 2006; Yu et al., 2007). For example, dermal fibroblasts can be reprogrammed to a pluripotent state by ectopic expression of a cocktail of pluripotent factors including Oct-4 (POU5F1), Sox-2, Klf-4, c-Myc, Nanog, and Lin28 (Takahashi et al., 2007; Yu et al., 2007), With the exception of Oct4, further studies indicated that the majority of these factors could be eliminated by use of unique stem/progenitor cells (Heng et al.; Aasen et al. 2008; Eminli et al. 2008; Eminli et al. 2009; Kim et al. 2009) or, alternatively, by addition of chemicals targeting the epigenome of dermal fibroblast sources (Shi et al. 2008; Lyssiotis et al. 2009). These studies demonstrate there are several approaches and methods for generation of iPSCs, however, the cellular and molecular mechanisms underlying reprogramming to the pluripotent state remain largely unknown (Jaenisch and Young, 2008). Although iPSCs can be differentiated towards the blood fate, the resulting hematopoietic cells preferentially generate primitive blood cells that utilize embryonic programs. Moreover, the methods remain inefficient, making it difficult to contemplate transplantation or modeling hematological diseases (Lengerke and Daley, 2010). Characterization of these processes is further complicated by cellular intermediates that fail to establish a stable pluripotent state, potentially due to the inability to achieve the correct combination, stoichiometry, or expression levels of reprogramming factors ideal for complete pluripotency induction (Chan et al., 2009; Kanawaty and Henderson, 2009; Lin et al., 2009; Mikkelsen et al., 2008). Consistent with this idea, intermediate cells derived from fibroblasts have been shown to co-express genes associated with several differentiated lineages (neurons, epidermis, and mesoderm) (Kanawaty and Henderson, 2009; Mikkelsen et al., 2008), nevertheless the exact identity and differentiation potential of these cell types remain elusive. This creates the possibility that under unique conditions the fibroblasts expressing a small subset of transcription factors can be induced to differentiate towards specified lineages without achieving pluripotency, as recently been demonstrated by converting fibroblasts into specific cell types such as neurons, cardiomyocytes, and macrophage-like cells (Feng et al., 2008; Ieda et al., 2010; Vierbuchen et al., 2010). While these studies have examined fibroblast conversion in the murine model, this concept remains to be extrapolated for human applications.
Previous studies have shown that proteins containing POU domains, such as Oct-4, along with Oct-2 (POU2F2) and Oct-1 (POU2F1) bind similar DNA target motifs (Kang et al., 2009). Whilst both Oct-2 and Oct-1 play a role in hematopoietic development (Brunner et al., 2003; Emslie et al., 2008; Pfisterer et al., 1996), Oct-4 is yet to be implicated in this process. Nonetheless, recent studies have predicted that Oct-4 possesses the capacity to bind to the promoters of the hematopoietic genes Runx1 and CD45, thus potentially regulating their expression (Kwon et al., 2006; Sridharan et al., 2009). Despite the similarities in binding and regulation, the exact functional role of individual Oct family members appears to be cell context specific (Kang et al., 2009; Pardo et al., 2010).
The ability to generate pluripotent stem cells from human dermal fibroblasts allows for generation of complex genetic disease models, and provides an unprecedented source for autologous transplantation without concern of immune rejection (Takahashi and Yamanaka 2006; Hanna et al. 2007; Yu et al. 2007; Okita et al. 2008; Park et al. 2008; Park et al. 2008b; Soldner et al. 2009).
Although a variety of somatic cell types can be reprogrammed, the vast majority of studies aimed at characterizing the mechanisms that govern the reprogramming process utilize fibroblasts (Takahashi and Yamanaka 2006; Takahashi et al. 2007; Wernig et al. 2007; Yu et al. 2007; Aoi et al. 2008; Brambrink et al. 2008; Eminli et al. 2008; Hanna et al. 2008; Huangfu et al. 2008; Lowry et al. 2008; Stadtfeld et al. 2008; Zhou et al. 2008; Carey et al. 2009; Feng et al. 2009; Gonzalez et al. 2009; Guo et al. 2009; Kaji et al. 2009; Utikal et al. 2009; Woltjen et al. 2009; Yusa et al. 2009; Zhou et al. 2009). As such, the current understanding of the molecular mechanisms and cellular nature of reprogramming is nearly exclusively derived from fibroblast-based reprogramming. Fibroblasts can be generated from multiple tissue sites including dermal skin, however, little is known about the origins and composition of fibroblasts used experimentally.
Cellular reprogramming to the pluripotent state was originally demonstrated using in vitro cultured mammalian fibroblasts (Takahashi and Yamanaka 2006). To date, iPSCs have been derived from a number of other tissue-derived cells including liver, pancreas, intestine, stomach, adipose, melanocytes, and hematopoietic sources (Aoi et al. 2008; Hanna et al. 2008; Zhou et al. 2008; Eminli et al. 2009; Sun et al. 2009; Utikal et al. 2009) using a variety of transcription factors including the oncogenes c-myc and klf4 (Takahashi and Yamanaka 2006; Takahashi et al. 2007; Aasen et al. 2008; Hanna et al. 2008; Park et al. 2008; Eminli et al. 2009; Hanna et al. 2009; Woltjen et al. 2009; Zhao et al. 2009). To date, the reprogramming process remains inefficient, but can be enhanced by utilization of initial cell types that already possess stem/progenitor proliferative capacity (Kim et al. 2009; Eminli et al. 2008; Eminli et al. 2009), or by enhancing cell cycle state by knocking down inhibitors of cell cycle progression such as p53/p21 (Kawamura et al. 2009; Li et al. 2009; Utikal et al. 2009). However, altering cell cycle regulators or introduction of oncogenes increases the risk of uncontrolled growth and tumor formation and thus raises potential safety concerns for future human therapeutic applications (Lebofsky and Walter 2007; Okita et al. 2007; Nakagawa et al. 2008; Markoulaki et al. 2009).